Fig 1: Rosiglitazone increases apoptosis and autophagy in GH3 cells(A) SW033291 significantly reversed the growth inhibitory effect of rosiglitazone on GH3 cells. GH3 cells were treated with rosiglitazone (Ros, 50 µM) or rosiglitazone (Ros, 50 µM) plus SW033291 (500 nM) (Ros + SW033291) for 24, 48, 72, or 96 hr. Cells were added with DMSO as control (Ctrl). GH3 cell proliferation was assessed using the CCK8 kit. Data were represented as mean ± SD. ** denotes p < 0.01 versus Ctrl. ## denotes p < 0.01 versus Ros.(B) SW033291 significantly abolished the increased both early and late apoptosis of GH3 cells induced by rosiglitazone. GH3 cells were treated with rosiglitazone (Ros, 50 µM) or rosiglitazone (Ros, 50 µM) plus SW033291 (500 nM) (Ros + SW033291) for 24 hr. Cells were added with DMSO as control (Ctrl). Cell apoptosis (both early apoptosis and late apoptosis) was assessed by Annexin V/propidium iodide (PI) assay. Data were represented as mean ± SD. ** denotes p < 0.01 versus Ctrl. ## denotes p < 0.01 versus Ros.(C) SW033291 impeded the increased cleaved caspase 3 protein expression caused by rosiglitazone in GH3 cells. GH3 cells were treated with rosiglitazone (Ros, 50 µM) or rosiglitazone (Ros, 50 µM) plus SW033291 (500 nM) (Ros + SW033291) for 24 hr. Cells were added with DMSO as control (Ctrl). The protein level of cleaved caspase 3 was assessed by western blot.(D) SW033291 reversed the rosiglitazone-induced changes of several proteins involved in autophagy pathways. GH3 cells were treated with rosiglitazone (Ros, 50 µM) or rosiglitazone (Ros, 50 µM) plus SW033291 (500 nM) (Ros + SW033291) for 24 hr. Cells were added with DMSO as control (Ctrl). The protein levels of Beclin1 (BECN1), ATG5, ATG7, p62, mammalian target of rapamycin (mTOR), p-mTOR, Akt, p-Akt, p-EIF4EBP1, and EIF4EBP1 were assessed by western blot.(E) SW033291 reversed the increased autophagy in the rosiglitazone-treated GH3 cells. Confocal analysis of GH3 cells expressing tandem mCherry-GFP-LC3 treated with treated with rosiglitazone (Ros, 50 µM) or rosiglitazone (Ros, 50 µM) plus SW033291 (500 nM) (Ros + SW033291) for 24 hr. Cells were added with DMSO as control (Ctrl). Scale bar represents 10 µm.